Mammalian skin and its appendages function as the outermost barrier of the body to protect inner organs from environmental hazards and keep essential fluids within the body. During embryonic development, a single layer of epidermal skin stem cells gives rise to the epidermis, hair follicles (HF) and the sebaceous gland, a process orchestrated by an array of regulatory pathways. In adult, homeostasis of each of three skin lineages is maintained long-term through self-renewal and differentiation of distinct skin stem cells. Through extensive investigation, much has been learned about the regulatory networks that control skin morphogenesis during embryonic development and self-renewal and differentiation of adult skin stem/progenitor cells. Recently, however, a novel layer of regulation mediated by miRNAs has been implicated in animal development. MiRNAs are a family of non-coding, small RNAs (~19-24nt) expressed in a wide range of animals and plants. Mature miRNAs act by specifically coupling with their target mRNAs at the 3' untranslated region to regulate the translation and/or mRNA stability. It is estimated that more than one third of protein-encoding mRNAs are regulated by miRNAs. In turn, miRNA-mediated regulation is believed to have a widespread impact on both protein output of transcriptome and evolution of gene regulatory networks. MiRNAs' potentials in globally regulating gene expression and developmental transitions during mammalian skin morphogenesis as well as their potential involvement in human skin diseases have attracted significant interests in the role of these novel regulators in skin biology. This proposal is based on my earlier findings and built to address three outstanding questions of miRNA biology and skin biology by focusing on miR-203, the most abundantly and specifically expressed miRNA in mammalian skin. In aim 1, I will dissect the underlying mechanisms of miR-203's functions by addressing an important question, that is, how each miRNA target contributes to the miRNA's functions in cell cycle regulation. In aim 2, I will investigate how the regulation of miR-203 itself is achieved in the skin by exploring its transcriptional regulation. In aim 3, I will begin to investigate miR-203's in vivo functions by examining its physiological functions during the earliest basal-suprabasal transition around E14 with a constitutive knockout mouse model. Taken together, studies proposed here, if successful, will significantly enhance our knowledge about individual miRNA's expression, function and regulation in the skin. The knowledge gained from these studies under normal development condition will also pave the way to investigate miRNA's roles in human diseases e.g. skin cancers where regulatory networks go awry. PUBLIC HEALTH RELEVANCE: MiRNA is a class of newly discovered regulatory genes. Recent studies have clearly demonstrated its critical functions in mammalian development and disease progression. In this proposal, I will investigate their functions in mammalian skin development. Knowledge gained in this research will lay a foundation to examine their functions in human skin diseases.